CN104450893B - Probe group and gene chip for detecting bladder cancers - Google Patents

Abstract

The invention discloses a probe group for detecting bladder cancers. The probe group comprises 25 probes 1-25, wherein the nucleotide sequences of the 25 probes 1-25 sequentially are sequences 1-25 in a sequence table; the oligonucleotide probes 1-25 are cured on a vector material by virtue of arm molecules to form a probe array, namely a gene chip. When applied to the early detection of the bladder cancers, the probe group and the gene chip particularly have the advantages that (1) the specificities are high, namely 25 indexes of pathogen of the bladder cancer can be simultaneously detected by virtue of a new gene combination for detecting the bladder cancers, and the specificity reach up to 95%; (2) the sensitivities and the accuracies are high, namely the sensibilities reach 95%, and the clinic coincidence rates reach 95%; (3) the bladder cancers can be rapidly detected and prognostically monitored; (4) the probe group and the gene chip can be used for diagnosing and prognostically monitoring various bladder cancers and are particularly used for diagnosing early-phase bladder cancer.

Description

A kind of probe groups for detecting bladder cancer and gene chip

Technical field

The present invention relates to field of biological genes, be specifically related to a kind of probe groups for detecting bladder cancer and gene chip.

Background technology

Bladder cancer refers to the malignant tumor in bladder mucosa.It is the modal malignant tumor of urinary system, is also whole body ten One of big kinds of tumor.Account for first of China's Genitourinary system sickness rate, and its sickness rate is only second to prostate in west Cancer, occupies the 2nd.The sickness rate of national tumor registration area bladder cancer in 2012 is 6.61/10 ten thousand, row Cancer Mortality 9th.Bladder cancer can betide any age, even child.Its sickness rate with the age increase and increase, the age 50 occurred frequently～ 70 years old.Male bladder cancer morbidity is 3～4 times of women.Bladder cancer cancer mainly according to patient's history of past illness, family history, in conjunction with Symptom makes preliminary judgement with having a medical check-up, and carries out coherence check further.Inspection method includes routine urianlysis, Urine exfoliative cell Learn, urinate the inspections such as tumor marker, abdominal part and Visual quality.Decide whether that row cystoscope, vein are urinated according to above-mentioned inspection result Road radiography, pelvic cavity CT are or/and the inspection such as pelvic MRI is clarified a diagnosis.Wherein, cystoscopy is the main side of diagnosing bladder cancer Method.At present, bladder cancer does not still have effective early diagnosis and prognostic indicator.

Gene chip (Gene Chip) is that substantial amounts of oligonucleotide molecules is fixed on solid phase carrier formation DNA microarray, borrows The sequence information of DNA sample is efficiently understood and analyzes by the characteristic helping nucleic acid molecule hybridization pairing.

Gene chip originates from the eighties in 20th century, has obtained swift and violent development in recent years, but has mainly been applied to medicine and research Field, is not the most applied to the report of bladder cancer detection in early days by biochip technology.

Summary of the invention

The purpose of the present invention is aiming at above-mentioned defect of the prior art, it is provided that specificity is good, highly sensitive, accuracy rate is high For detecting probe groups and the gene chip of bladder cancer.

To achieve these goals, the technical scheme that the present invention provides is: a kind of probe groups for detecting bladder cancer, by probe 1-probe 25 totally 25 probe compositions, the sequence that the nucleotide sequence of described probe 1-probe 25 respectively is in sequence table 1-25。

For detecting the probe groups of bladder cancer disease, including 25 genetic fragments, wherein the 25th kind of genetic fragment is Quality Control gene, 25 genetic fragments are to design for 25 indexs of bladder cancer, utilize 25 genetic fragments jointly to detect bladder cancer base Because of the change of express spectra, and then bladder cancer is carried out detection and prognostic monitoring.

As shown in Table 1 and Table 2, wherein table 1 is sequence 1-13 to 25 kinds of genetic fragments, and table 2 is sequence 14-25:

Table 1

Table 2

The detailed sequence of 25 genetic fragments is as shown in sequence 1-25 in sequence table 1-2.

The above-mentioned probe groups for detecting bladder cancer can be applicable to biochip technology and (includes biochip technology, protein chip skill Art, tissue array technology, cell chip technology etc.), polymerase chain reaction (PCR), hybridization and its His pathogen gene diagnostic techniques.

Second object of the present invention there is provided a kind of gene chip for detecting bladder cancer, and described gene chip is by above-mentioned Oligonucleotide probe 1-25 solidified the probe array formed on a support material by arm molecule.

Further, above-mentioned a kind of gene chip for detecting bladder cancer, described probe 1-probe 24 is detection oligonucleoside Acid probe, probe 25 is quality control oligonucleotide probe.

Further, above-mentioned a kind of gene chip for detecting bladder cancer, the probe labeling techniques in described gene chip is Isotope labeling, biotin labeling, digoxigenin labeled, fluorescein labelling, CI labelling, colloid gold label or combined mark.

Third object of the present invention there is provided the above-mentioned gene chip for detecting bladder cancer in detection in early days bladder cancer Application.

Further, above-mentioned application, during described detection bladder cancer in early days, detection sample is bladder cancer flesh tissue or wing The Urine exfoliative cell of Guang cancer patient.

The invention have the benefit that the early stage detection that probe groups and gene chip are applied to bladder cancer provided by the present invention exists China still belongs to the first time, and this will be greatly facilitated the development of bladder cancer early diagnosis, specifically has the advantage that

1, specificity is good: the diagnosis of existing bladder cancer is only to detect one or several index to lack specificity, so easily drawing Playing mistaken diagnosis, the misdiagnosis rate of bladder cancer is more up to more than 70% especially in early days.The present invention is for detecting the new gene of bladder cancer Combination can detect 25 indexs of bladder cancer pathogen simultaneously, and specificity is up to 95%；

2, sensitivity and accuracy rate are high: sensitivity reaches 95%, and clinical coincidence rate (accuracy rate) reaches 95%；

3, bladder cancer can be carried out quick detection and prognostic monitoring；

4, can be used for diagnosis and the prognostic monitoring of various types of bladder cancer, be used especially for the diagnosis of bladder cancer in early days.

For ensureing the quality of gene chip, it should be noted that following item:

1) extraction of sample nucleic: the nucleic acid of extraction must have higher purity (OD260/0D280 ratio should be greater than 1.50) and A certain amount (answers 10ng-100ng), to ensure quality and the specific amplification of labelling, if purity is inadequate, after may causing There is non-specific point in the hybridization in face；

2) labelling of sample nucleic: the label of incorporation must reach certain concentration, and (label that each reaction mixes is 0.1-1 μ g), to ensure the sensitivity of hybridization point, if concentration is inadequate, results of hybridization often there will be false negative；

3) homologous nucleic acid hybridization: should grasp hybridization temperature and control at about 42 DEG C, temperature is too low, it may appear that Non-specific hybridization point, Making specificity reduce, temperature is too high, false negative the most often occurs.

The method of bladder cancer early diagnosis gene chip and conventional sense disease has the advantage that to comparing

1, accurate, gene chip is provided with the positive and negative control, can avoid the wrong diagnosis that environment and anthropic factor cause；

2, quick, conventional sense needs the time of one week, and chip only needs about 8 hours；

3, sensitive, the susceptiveness (0.1pg) of chip is high 1000 times than round pcr；

4: the PCR that contains much information the most only examines a kind of index, and chip once can detect 20～30 kind of index.

Accompanying drawing explanation

Fig. 1 is chip form design figure.

Fig. 2 is chip detection result.

Detailed description of the invention

Embodiment 1:

One, for detecting the preparation of the gene chip of the probe groups of bladder cancer:

1, material and method

1.1 materials:

1.1.1 reagent:

Main agents RNA extract reagent (Trizol Reagent), saturated phenol, DEPC, DNA marker, Oligod (T) 18, DIG DNA marker test kit, digoxin hybridization check test kit, be purchased from the raw work in Shanghai.Reverse transcriptase M-MLV (RNase H-), Taq enzyme, Agarose Gel, RNase inhibitor, agarose (Agarose), DNA purification kit, be purchased from Dalian Precious biotech firm.Primer is synthesized by Shanghai Sheng Gong bio-engineering corporation；Other reagent are domestic analytical pure.

1.1.2 clinical data:

(1) bladder cancer flesh tissue sample: tissue specimen takes from the People's Hospital of Gansu Province in the phase in April ,-2013 in October, 2011 Between the fresh bladder cancer specimen of excision, front all do not do chemotherapy or radiotherapy, Postoperative pathological confirms, simultaneously away from borderline tumor 5- More than 10cm cuts normal structure (verified by postoperative pathology cancer-free cell invades profit) as comparison.I.e. put in specimen rear 5min in vitro In liquid nitrogen container frozen.

(2) bladder cancer patients Urine exfoliative cell sample: detection sample is that the People's Hospital of Gansu Province is in April ,-2013 in April, 2012 The 100 example cell detection samples (wherein bladder cancer sample 50 example) of period bladder cancer and normal person's Urine exfoliative cell sample are used respectively Chip method carries out comparison and detection, is stored in-80 DEG C.

1.1.3 instrument and equipment:

Instrument and equipment: PCR instrument is purchased from Hangzhou Lang Ji；Sample applicator；Hybridization instrument；Scanner is designed, designed, entrusts and produces Producer manufactures.

1.2 design of primers:

Determine several sequences of bladder cancer tumor markers, then use DNAStar to analyze and design these primers.Concrete primer is such as Shown in above-mentioned table 1-table 2.

The making of 1.3 gene chips:

1.3.1 the extraction (bladder cancer flesh tissue sample) of bladder cancer nucleic acid:

(1) from liquid nitrogen, take out frozen flesh tissue 200mg, the mortar of Liquid nitrogen precooler grinds tissue, uses Trizol Method extracts total tissue RNA, measures total serum IgE purity and by Oligo dT cellulose chromatography purified mRNA.

(2) use first chain of reverse transcriptase M-MLV reverse transcription synthesis cDNA, be used for preparing probe template.

1.3.2 the amplification of nucleic probe:

(1) nucleic acid obtained with 1.3.1 is as template, uses the primer designed by table 1-2, amplifies segment respectively, with As probe.Amplified fragments particular sequence is sequence shown in sequence table middle probe 1-probe 25.

(2) PCR reaction system 50.0 μ L, including 10 × Buffer5.0 μ L, 25mmol dNTPmixture0.5, TaqDNA enzyme 1.0 μ L, each 1.0 μ L of purpose primer upstream and downstream, cDNA template 1.0 μ L, tri-distilled water 40.5 μ L, reaction condition: 94 DEG C of denaturations 5min；94 DEG C of degeneration 30s, 55 DEG C of annealing 30s, 72 DEG C extend 60s, 30 circulations；Reaction is terminated after 72 DEG C of 10min.

(3) utilize small pieces segment DNA purification kit that the 25 kinds of fragment PCR product obtained are purified.

1.3.3 some film:

By micro-sampling device by the probe points that obtained on the nylon membrane of pretreatment.

According to the figure of chip form design shown in Fig. 1, carry out chip point sample with manual point sample instrument.Operation sequence:

1) GAPDH, the sampling liquid of 25 kinds of genes such as ACTIN, CYCLIND1, at 100 DEG C of degeneration 5min, puts rapidly ice, ice-cold 10min Above；

2) nylon membrane (purchased from Amersco company, positively charged) sterilizing distilled water soaks 10min, unsettled dries；

3) every 1 μ l is above-mentioned probe fragment point (every nylon membrane area about 20cm on nylon membrane²)；

4) wet film is according to 5min under uviol lamp, takes out and allows film natural drying.

The process of 1.4 samples:

1.4.1 the extraction (bladder cancer patients Urine exfoliative cell sample) of nucleic acid:

1) the same 1.3.1 of bladder cancer flesh tissue Sample Method, uses primer 5 ,-AAG CAG TGG TAA CAA CGC AGA GTA CGC GGG-3 ' and 5 ,-AAG CAG TGG TAA CAA CGC AGA GTA CT (30) n-1n-3, reverse transcription synthesis cDNA's First chain；

2) use Trizol method to extract bladder cancer patients uterine cavity exfoliative cyte sample total serum IgE, use primer 5 ,-AAG CAG TGG TAA CAA CGC AGA GTA CGC GGG-3 ' and 5 ,-AAG CAG TGG TAA CAA CGC AGA GTA CT (30) n-1n-3, First chain of reverse transcription synthesis cDNA.

1.4.2 the amplification of nucleic acid:

1) nucleic acid obtained with 1.4.1 is as template, uses the universal primer 5 of design ,-AAG CAG TGG TAA CAA CGC AGA GT-3 amplifies genetic fragment, using as detection sample；

2) PCR reaction system 25.0 μ L, including 10 × Buffer 2.5 μ L, 10mmol dNTPmixture 0.5 μ l, ground High octyl-11-dUTP 0.5 μ l, TaqDNA enzyme 0.5 μ L, universal primer upstream and downstream each 0.5 μ L, cDNA template 5.0 μ L, Tri-distilled water 15 μ L, reaction condition: 94 DEG C of denaturations 5min；94 DEG C of denaturations 30s, 65 DEG C of annealing 30s, 68 DEG C of extensions 500s, 30 circulations；Reaction is terminated after 72 DEG C of 10min.

1.5 hybridization:

Adding 500 μ l prehybridization solutions on gene chip, 42 DEG C process 40min, take the product of 20 μ l1.4.2 in 1.5ml from In heart pipe, 10min.-20 DEG C of cooling 5min of 100 DEG C of degeneration, add hybridization solution 450 μ l, mixing, on gene chip, 42 DEG C of hybridization 1h.Respectively with 2 × SSC, 0.1 × SDS and 0.1 × SSC, 0.1 × SDS respectively wash three times, add the BSA of 400 μ l3%, 42 DEG C of closings 30min, adds 1.6 μ l AV-AP, 42 DEG C of enzyme connection 20min, first washes film three times with cleaning mixture, add colour developing afterwards in 450 μ l buffer Liquid (450 μ l substrate solution+1.6 μ l BCIP+1.6 μ l NBT), room temperature or 42 DEG C of colour developings 3～5min.With distilled water or tap water Rinse, color development stopping.

The analysis of 1.6 results of hybridization:

It is scanned analyzing by hybridizing the homemade chip scanner of the complete DNA array dried, according to the analysis of a large amount of samples, Arranging reading less than 0.05 is feminine gender, and more than 0.1 is positive, is suspicious between the two.Print diagnosis report.

Sample carries out PCR or RT-PCR with other detection methods to compare simultaneously, and result is as shown in table 3.

Table 3

The positive and negative control: be divided into the GAPDH of people, ACTIN, CYCLIND1 genetic fragment, add response system, to monitor labelling With hybridization, Color Appearance System the most effective.

2, result:

The result of 2.1 accuracy rate detections:

The most respectively to GAPDH, 25 kinds of genes such as ACTIN, CYCLIND1 are detected, results of hybridization: 30 chips divide GAPDH, 25 kinds of gene specific hybridization points such as ACTIN, CYCLIND1, testing result such as Fig. 2 do not occur, does not has non-specific miscellaneous The appearance of intersection point.

The testing result of 2.2 recall rates:

Utilize bladder cancer gene diagnosis chip that the Urine exfoliative cell of 50 example patients is detected, find 48 example test positive, 1 Example is the weak positive, and 1 example is negative；Its specificity reaches 95%, sensitivity reaches 95%, clinical coincidence rate (accuracy rate) reaches 95%, base Reach on Ben to design requirement.

Finally it is noted that the foregoing is only the preferred embodiments of the present invention, it is not limited to the present invention, although Being described in detail the present invention with reference to previous embodiment, for a person skilled in the art, it still can be to front State the technical scheme described in each embodiment to modify, or wherein portion of techniques feature is carried out equivalent.All at this Within bright spirit and principle, any modification, equivalent substitution and improvement etc. made, should be included in protection scope of the present invention Within.

Sequence table

<110>Lanzhou Baiyuan Gene Technology Co., Ltd.

<120>a kind of probe groups for detecting bladder cancer and gene chip

<210> 1

<211> 175

<212> DNA

<213> MCM4

<400> 1

cacggctcaa tgtaggctta acctcctggg ctcaggtgta tgtcactatg cccggctact 60

ttttgtattt tttggtagag acggcttcgc cacgttgccc aggctgcaag cgatatgcct 120

aggctcaagc gatctgccca cctcaacttc cggaagtgct gagattacag gtgtg 175

<210> 1

<211> 223

<212> DNA

<213> CDK1

<400> 2

actgctcgca cttggcttca aagctggctc ttggaaattg agcggagagc gacgcggttg 60

ttgtagctgc cgctgcggcc gccgcggaat aataagccgg gatctaccat acccattgac 120

taactatgga agattatacc aaaatagaga aaattggaga aggtacctat ggagttgtgt 180

ataagggtag acacaaaact acaggtcaag tggtagccat gaa 223

<210> 1

<211> 197

<212> DNA

<213> SFRP1

<400> 3

tgtaatccag tcggcttgtt cttgcagcat tcccgctccc ttccctccat agccacgctc 60

caaaccccag ggtagccatg gccgggtaaa gcaagggcca tttagattag gaaggttttt 120

aagatccgca atgtggagca gcagccactg cacaggagga ggtgacaaac catttccaac 180

agcaacacag ccactaa 197

<210> 1

<211> 197

<212> DNA

<213> UBE2C

<400> 4

cagtcgtgtt ctccgagttc ctgtctctct gccaacgccg cccggatggc ttcccaaaac 60

cgcgacccag ccgccactag cgtcgccgcc gcccgtaaag gagctgagcc gagcgggggc 120

gccgcccggg gtccggtggg caaaaggcta cagcaggagc tgatgaccct catgatgtct 180

ggcgataaag ggatttc 197

<210> 1

<211> 250

<212> DNA

<213> IL6

<400> 5

gtacatcctc gacggcatct cagccctgag aaaggagaca tgtaacaaga gtaacatgtg 60

tgaaagcagc aaagaggcac tggcagaaaa caacctgaac cttccaaaga tggctgaaaa 120

agatggatgc ttccaatctg gattcaatga ggagacttgc ctggtgaaaa tcatcactgg 180

tcttttggag tttgaggtat acctagagta cctccagaac agatttgaga gtagtgagga 240

acaagccaga 250

<210> 1

<211> 195

<212> DNA

<213> GHR

<400> 6

gctcggaggt cctacaggta tggatctctg gcagctgctg ttgaccttgg cactggcagg 60

atcaagtgat gctttttctg gaagtgaggc cacagcagct atccttagca gagcaccctg 120

gagtctgcaa agtgttaatc caggcctaaa gacaaattct tctaaggagc ctaaattcac 180

caagtgccgt tcacc 195

<210> 1

<211> 189

<212> DNA

<213> FGFR1

<400> 7

cagaggatgg tgaggcgaag gccaggttgg gggcagtgtt gtggccctgg ggcccagccc 60

caaactgggg gctctgtata tagctatgaa gaaaacacaa agtgtataaa tctgagtata 120

tatttacatg tctttttaaa agggtcgtta ccagagattt acccatcggg taagatgctc 180

ctggtggct 189

<210> 1

<211> 246

<212> DNA

<213> CXCL10

<400> 8

gaacctccag tctcagcacc atgaatcaaa ctgccattct gatttgctgc cttatctttc 60

tgactctaag tggcattcaa ggagtacctc tctctagaac tgtacgctgt acctgcatca 120

gcattagtaa tcaacctgtt aatccaaggt ctttagaaaa acttgaaatt attcctgcaa 180

gccaattttg tccacgtgtt gagatcattg ctacaatgaa aaagaagggt gagaagagat 240

gtctga 246

<210> 1

<211> 176

<212> DNA

<213> FGF13

<400> 9

ggtctgcgag tggtggctat ccaaggagtt caaaccaagc tgtacttggc aatgaacagt 60

gagggatact tgtacacctc ggaacttttc acacctgagt gcaaattcaa agaatcagtg 120

tttgaaaatt attatgtgac atattcatca atgatatacc gtcagcagca gtcagg 176

<210> 1

<211> 198

<212> DNA

<213> PTCH1

<400> 10

ctcgcctatg cctgtctaac catgctgcgc tgggactgct ccaagtccca gggtgccgtg 60

gggctggctg gcgtcctgct ggttgcactg tcagtggctg caggactggg cctgtgctca 120

ttgatcggaa tttcctttaa cgctgcaaca actcaggttt tgccatttct cgctcttggt 180

gttggtgtgg atgatgtt 198

<210> 1

<211> 240

<212> DNA

<213> GSTM3

<400> 11

atagtgtgag ggagcggtat gctctaccct tgtgtattta aggcaggaca gagaaattga 60

ggatgcccct ggggctagat cgatgatatg accagaaatc aaaaagggaa tgcattattt 120

attgctgtgt aacactgtta agagaggagg tagttaaggg atggataggc acatagagag 180

ggaaggtctc aggagaaggg atagaaagga actatgttca caagaacaac ggcaagacag 240

<210> 1

<211> 223

<212> DNA

<213> ADH1B

<400> 12

tcgtggagag tgttggagaa ggggtgacta cagtcaaacc aggtgataaa gtcatcccgc 60

tctttactcc tcagtgtgga aaatgcagag tttgtaaaaa cccggagagc aactactgct 120

tgaaaaatga tctaggcaat cctcggggga ccctgcagga tggcaccagg aggttcacct 180

gcagggggaa gcccattcac cacttccttg gcaccagcac ctt 223

<210> 1

<211> 151

<212> DNA

<213> AKR1C1

<400> 13

aagccaggtg aggaagtgat cccaaaagat gaaaatggaa aaatactatt tgacacagtg 60

gatctctgtg ccacatggga ggccgtggag aagtgtaaag atgcaggatt ggccaagtcc 120

atcggggtgt ccaacttcaa ccgcaggcag c 151

<210> 1

<211> 196

<212> DNA

<213> CNTN1

<400> 14

tgtggttgct ggtcagtcat cttgtgataa tatctattac tacctgttta gcagagttta 60

catggtatag aagatatggt catggagttt ctgaggaaga caaaggattt ggaccaattt 120

ttgaagagca gccaatcaat accatttatc cagaggaatc actggaagga aaagtctcac 180

tcaactgtag ggcacg 196

<210> 1

<211> 217

<212> DNA

<213> CCND2

<400> 15

gatgctggag gtctgtgagg aacagaagtg cgaagaagag gtcttccctc tggccatgaa 60

ttacctggac cgtttcttgg ctggggtccc gactccgaag tcccatctgc aactcctggg 120

tgctgtctgc atgttcctgg cctccaaact caaagagacc agcccgctga ccgcggagaa 180

gctgtgcatt tacaccgaca actccatcaa gcctcag 217

<210> 1

<211> 228

<212> DNA

<213> SDC2

<400> 16

agaggaggag aaggaggagg acccggggag ggaggcgcgg cgcgggagga ggaggggcgc 60

agccgcggag ccagtggccc cgcttggacg cgctgctctc cagatacccc cggagctcca 120

gccgcgcgga tcgcgcgctc ccgccgctct gcccctaaac ttctgccgta gctccctttc 180

aagccagcga atttattcct taaaaccaga aactgaacct cggcacgg 228

<210> 1

<211> 181

<212> DNA

<213> THBS1

<400> 17

gatggaattg gtgatgcctg tgatgatgac gatgacaatg ataaaattcc agatgacagg 60

gacaactgtc cattccatta caacccagct cagtatgact atgacagaga tgatgtggga 120

gaccgctgtg acaactgtcc ctacaaccac aacccagatc aggcagacac agacaacaat 180

g 181

<210> 1

<211> 238

<212> DNA

<213> MMP2

<400> 18

accacagcca actacgatga tgaccgcaag tggggcttct gccctgacca agggtacagc 60

ctgttcctcg tggcagccca cgagtttggc cacgccatgg ggctggagca ctcccaagac 120

cctggggccc tgatggcacc catttacacc tacaccaaga acttccgtct gtcccaggat 180

gacatcaagg gcattcagga gctctatggg gcctctcctg acattgacct tggcaccg 238

<210> 1

<211> 223

<212> DNA

<213> CDC42

<400> 19

tacgaccgct gagttatcca caaacagatg tatttctagt ctgtttttca gtggtctctc 60

catcttcatt tgaaaacgtg aaagaaaagt gggtgcctga gataactcac cactgtccaa 120

agactccttt cttgcttgtt gggactcaaa ttgatctcag agatgacccc tctactattg 180

agaaacttgc caagaacaaa cagaagccta tcactccaga gac 223

<210> 1

<211> 231

<212> DNA

<213> DAPK1

<400> 20

cgtgaacatc aagaaccgag aaggagagac gcccctcctg acagcctctg ccaggggcta 60

ccacgacatc gtggagtgtc tggccgaaca tggagccgac cttaatgctt gcgacaagga 120

cggacacatt gcccttcatc tggctgtaag acggtgtcag atggaggtaa tcaagactct 180

cctcagccaa gggtgtttcg tcgattatca agacaggcac ggcaatactc c 231

<210> 1

<211> 238

<212> DNA

<213> STMN1

<400> 21

gaaacgagag cacgagaaag aagtgcttca gaaggcaata gaagagaaca acaacttcag 60

taaaatggca gaagagaaac tgacccacaa aatggaagct aataaagaga accgagaggc 120

acaaatggct gccaaactgg aacgtttgcg agagaagatg tacttctgga ctcacgggcc 180

tggggcccac ccagcacaga tctctgctga gcaatcttgt ctccactctg ttcctgcc 238

<210> 1

<211> 158

<212> DNA

<213> IGFBP3

<400> 22

ccagtagtca gcaaagagca gtttgaattt tcttgtcgct tcctatcaaa atattcagag 60

actcgagcac agcacccaga cttcatgcgc ccgtggaatg ctcaccacat gttggtcgaa 120

gcggccgacc actgactttg tgacttaggc ggctgtgt 158

<210> 1

<211> 210

<212> DNA

<213> IGF-I

<400> 23

tctctgaatc ttggctgctg gagccattca ttcagcaacc ttgtctaagt ggtttatgaa 60

ttgtttcctt atttgcactt ctttctacac aactcgggct gtttgtttta cagtgtctga 120

taatcttgtt agtctatacc caccacctcc cttcataacc tttatatttg ccgaatttgg 180

cctcctcaaa agcagcagca agtcgtcaag 210

<210> 1

<211> 193

<212> DNA

<213> VEGFA

<400> 24

ggcgaagaga agagacacat tgttggaaga agcagcccat gacagctccc cttcctggga 60

ctcgccctca tcctcttcct gctccccttc ctggggtgca gcctaaaagg acctatgtcc 120

tcacaccatt gaaaccacta gttctgtccc cccaggagac ctggttgtgt gtgtgtgagt 180

ggttgacctt cct 193

<210> 1

<211> 138

<212> DNA

<213> GAPDH

<400> 25

gcaccgtcaa ggctgagaac gggaagcttg tcatcaatgg aaatcccatc accatcttcc 60

aggagcgaga tccctccaaa atcaagtggg gcgatgctgg cgctgagtac gtcgtggagt 120

ccactggcgt cttcacca 138

Claims (4)

1. the probe groups being used for detecting bladder cancer, it is characterised in that be made up of probe 1-probe 25 totally 25 probes, sequence 1-25 that the nucleotide sequence of described probe 1-probe 25 respectively is in sequence table.

2. the gene chip being used for detecting bladder cancer, it is characterised in that described gene chip is that the probe 1-25 described in claim 1 is solidified the probe array formed on a support material by arm molecule.

A kind of gene chip for detecting bladder cancer the most according to claim 2, it is characterised in that described probe 1-probe 24 is detection oligonucleotide probe, and probe 25 is quality control oligonucleotide probe.

A kind of gene chip for detecting bladder cancer the most according to claim 3, it is characterized in that, the probe labeling techniques in described gene chip is isotope labeling, biotin labeling, digoxigenin labeled, fluorescein labelling, CI labelling, colloid gold label or combined mark.